Background. Natural killer (NK) cells eliminate incipient cancers and hematogenous metastases. NK mediated killing is inhibited y target cell HLA class i molecules, possibly through diverse HLA receptors recognize related HLA class I alleles, probably by contracting relatively conserved alpha-helical sites. Each NK cell expresses several crossreactive HLA receptors and recognizes multiple host HLA molecules. Recognition of target cell HLA inhibits NK mediated killing, proliferations, and cytokine production. Preliminary Studies. HLA-B*0702 protects transfected target cells from peripheral blood mononuclear cell )PBMC) NK mediated killing. Significantly increased killing is allowed by mutations in the B*0702 peptide binding groove, including the B pocket, and in the solvent- accessible T cell receptor contact site. The same mutations affect killing mediated by cytolytic T lymphocytes. HPLC separated peptides eluted from B*0702+ cells inhibit NK mediated killing of B*0702+ peptide transport deficient T2 cells. Our preliminary studies suggest that NK cells contact HLA-bound peptides and nearby HLA alpha-helical residues. Experimental Plan. We will test additions B*0702 mutants with PBMC NK cells nd NK clones to further refine the NK MHC contact site. We will examine how PBMC NK cells bearing the NKB1 putative HLA-B receptor and NK clones distinguish HLA alleles in three model systems; B*5101-B*0702, B*2705-B*0702, and Cw*0301-Cw*0601. If different HLA molecules inhibit NK cells in a quantitatively similar fashion, then expression of one, two, or three HLA genes will have an additive effect on NK mediated killing. We have devised several experimental approaches to identify HLA-binding peptides that inhibit NK cells. Peptide deficient T2 or acid stripped target cells will be incubated with HLA-binding synthetic or cellular peptides and tested in NK mediated killing assays. Alternatively, T2 cells will be transfected with individual minigenes or minigene libraries encoding HLA-binding peptides. Once protective peptides are identified, alanine substituted synthetic peptides will be tested for HLA binding and inhibition of K cells, allowing us to deduce NK receptor contact sites. Furthermore, we will t est if NK mediated killing, proliferations, and interferon gamma production are coordinately regulated by HLA and peptide variants. Whenever feasible, we will use both lymphoid and transformed oral keratinocyte target cells. Significance to Oral Cancer. Due to "field cancerization", oral cancer patients frequently suffer second malignancies and would benefit from enhanced NK surveillance. Identification of HLA structures that control NK mediated killing will help characterize newly described NK receptors. Identification of HLA or peptide variants that produce a 'split response" separating NK mediated killing, proliferation, and cytokine production, may suggest better ways to manipulate Nk attack on oral cancers. Protective HLA-binding peptides could inhibit NK attack of normal cells but not cancer cells.